论文部分内容阅读
采用玻璃针分离法，通过显微操作器成功地分离到大豆（ＧｌｙｃｉｎｅｍａｘＬ．）单染色体。将分离到的两条大豆染色体分别放入两个０．５ｍＬＥｐｐｅｎｄｏｒｆ管中，经Ｓａｕ３Ａ酶切，并在染色体ＤＮＡ片段两端加上Ｓａｕ３Ａ人工接头后，进行两轮ＰＣＲ扩增，得到０．３～３ｋｂ之间的ＤＮＡ片段。Ｓｏｕｔｈｅｒｎ杂交表明，这些大豆单染色体扩增片段与大豆基因组ＤＮＡ之间有同源性，从而证明两条单染色体ＤＮＡ确实已被成功地扩增了，同时表明两条不同的大豆单染色体扩增产物存在一定的差异。在常规的倒置显微镜下对小型染色体进行了显微分离，为小型单染色体ＤＮＡ的体外扩增及微克隆奠定了基础。
Single-chromosome isolation of soybean (Glycine max L.) was successfully performed by a micromanipulator using glass needle separation. The two isolated soybean chromosomes were placed in two 0.5mLEppendorf tubes, Sau3A digested, and both ends of the chromosomal DNA fragment Sau3A artificial joint, two rounds of PCR amplification to give 0.3 ~ 3kb between the DNA fragments. Southern hybridization showed that there was homology between these soybean single-chromosome amplified fragments and soybean genomic DNA, thus proving that the two single-chromosome DNAs have actually been successfully amplified and that two different soybean single-chromosome amplification products There are some differences. Microdissection of small chromosomes under conventional inverted microscope laid the foundation for the in vitro amplification and microcoloning of small single-chromosome DNA.